The goal of this proposal is to understand allergic disease processes and, in so doing, provide the means for the development of new modalities of therapy. The in vitro portion of this work focuses upon isolated and, most often, purified human basophils and mast cells. A major hypothesis of this proposal is that understanding the biochemistries -- the signal transduction mechanisms -- by which these cells secrete preformed and newly generated mediators will eventually allow us to develop pharmacologic agonists which block this release process. Several biochemical processes will be studied, including protein kinase C activation and changes in intercellular calcium concentrations; the role of these processes will also be assessed with available pharmacologic tools to evaluate their relationship to function. We will continue experiments which suggest that agonists which increase intracellular cAMP levels inhibit the production of arachidonic acid metabolites (LTC and PGD) more effectively and via different pathways than they affect histamine release. Function will also be evaluated in conjunction with morphologic changes in collaboration with Dr. Ann Dvorak. It has recently been appreciated that human mast cells are heterogeneous in terms of which mediators are produced, which secretagogues are effective and in the release mechanisms described above. We will study these parameters using human lung, skin, gut, uterine and BAL mast cells and basophils. In vivo mast cells and basophils function in the context of other cells; a major goal of this proposal is to explore those interactions. We have described a cytokine(s) coming from a variety of human cells which causes IgE-dependent histamine release from basophils. These histamine releasing factors (HRFS) are found in man in late phase reactions and their use has revealed a functional heterogeneity of IgE -- only one type of IgE (IgE+) can interact with HRF. We hypothesize that these cytokines play a major role in inflammation and we would like to purify and clone a HRF, to define the biochemical basis of IgE heterogeneity and to develop immunoassays for HRF and IgE+. Finally, it is our goal to test the relevance of the above studies in vivo in man. For this purpose, we are characterizing the pattern of mediator release and cellular infiltration after antigen challenge of the human upper and lower airways, skin and conjunctiva. By the use of inhibitors and antagonists, it should be possible to define which mediators and cells are most critically involved in the pathogenesis of allergic inflammation.